Senescent cell antigen (SCA), an aging antigen, is a protein that appears on old cells and acts as a specific signal for the termination of that cell by initiating the binding of IgG autoantibody and subsequent removal by phagocytes (1-14). This appears to be a general physiologic process for removing senescent and damaged cells in mammals and other vertebrates (4). Although the initial studies were done using human erythrocytes as a model, senescent cell antigen occurs on all cells examined (4). Besides its role in the removal of senescent and damaged cells, senescent cell antigen also appears to be involved in the removal of erythrocytes in clinical hemolytic anemias (7) and sickle cell anemia (8). It also appears to be involved in the removal of malaria-infected erythrocytes (15). Oxidation generates senescent cell antigen in situ (6).
Senescent cell antigen has been isolated from sialoglycoprotein mixtures with affinity columns prepared with IgG eluted from senescent cells (4). Both glycoprotein and protein stains of gels of the eluted material revealed a band migrating at a relative molecular weight of 62,000 in the component 4.5 region. These experiments suggested that the 62,000 M.sub.r glycoprotein carried the antigenic determinants recognized by IgG obtained from freshly isolated senescent cells. The 62,000 M.sub.r protein, but not the remaining sialoglycoprotein mixture from which it was isolated, abolished the phagocytosis-inducing ability of IgG eluted from senescent RBC in the erythrophagocytosis assay. This indicated that the 62,000 M.sub.r protein is the antigen which appeared on the membrane of the cells as they aged.
The aging antigen is derived from protein band 3, an important structural and membrane transport molecule (5). Band 3, is a ubiquitous protein (16-20). It has been found in diverse cell types and tissues besides erythrocytes, including hepatocytes (16), squamous epithelial cells (16), alveolar (lung) cells (16), lymphocytes (16), kidney (21), neurons (16, 17), and fibroblasts (16, 20). Band 3 is also present in nuclear (16), golgi (18), and mitochondrial membranes (19) as well as in cell membranes. Band 3-like proteins in nucleated cells participate in band 3 antibody induced cell surface patching and capping (16). Band 3 maintains acid-base balance by mediating the exchange of anions (e.g. chloride, bicarbonate) (22-24). Because of its central role in respiration of CO.sub.2, band 3 is the most heavily used ion transport system in vertebrate animals. Band 3 is a major transmembrane structural protein (25) which attaches the plasma membrane to the internal cell cytoskeleton by binding to band 2.1 (ankyrin) (26). The transport and the cytoskeletal domains can be separated by proteolysis with trypsin. Digestion with trypsin yields a 52,000 Da membrane bound transport domain and a 40,000 Da water-soluble cytoplasmic domain that binds cytoskeletal proteins. The transport domain of band 3 is highly conserved evolutionarily and no polymorphisms of it have been found. Senescent cell antigen (SCA) is generated on this transport domain (5).
Tanner et al. (Biochem. J. 256:703-712 (1988), incorporated herein by reference) isolated cDNA clones corresponding to the band 3 protein for human red blood cells and determined the deduced amino acid sequence. The paper discusses certain internal and external regions of the protein.
The inventors have determined the location and sequence of two peptide fragments corresponding to separate portions of the band 3 protein, each of which is immunoreactive with the IgG autoantibody and partially blocks the binding of such antibody to naturally occurring SCA. The inventors have also surprisingly discovered that the separate peptide fragments act together synergistically to substantially block the binding of the autoantibody to naturally occurring SCA and, therefore, together comprise a synthetic senescent cell antigen. The synthetic SCA and peptide components are expected to have a wide variety of scientific, diagnostic, and clinical applications.